Quantification of 5-Methylcytosine in DNA by the Chloroacetaldehyde Reaction
The study of changes in genome-wide levels of DNA methylation has become a key focus for understanding the epigenetic regulation of gene expression. Many procedures exist to study DNA methylation, falling into two categories: gene-specific and genome-wide. Genome-wide methylation analysis is best pe...
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| Format: | Article |
| Language: | English |
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Taylor & Francis Group
1999-10-01
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| Series: | BioTechniques |
| Online Access: | https://www.future-science.com/doi/10.2144/99274st05 |
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| author | Edward J. Oakeley Frédéric Schmitt Jean-Pierre Jost |
| author_facet | Edward J. Oakeley Frédéric Schmitt Jean-Pierre Jost |
| author_sort | Edward J. Oakeley |
| collection | DOAJ |
| description | The study of changes in genome-wide levels of DNA methylation has become a key focus for understanding the epigenetic regulation of gene expression. Many procedures exist to study DNA methylation, falling into two categories: gene-specific and genome-wide. Genome-wide methylation analysis is best performed by DNA hydrolysis followed by HPLC; however, it requires access to an HPLC machine, which is not always available. Alternative procedures, such as the radioactive labeling of CpG sites using SssI DNA methyltransferase, have been developed to address this problem, but it can only monitor CpG methylation changes, and CpNpG methylation is not detected. Here, we present a method for the analysis of DNA methylation in any sequence context by fluorescent labeling. We present control analyses using synthetic oligonucleotides of known methylation levels and a comparison of genomic DNA from two transgenic tobacco lines known to differ in their methylation levels. The results indicate that hygromycin-induced hypermethylation acts equally on all classes of methylatable cytosine, perhaps indicating a common mechanism. |
| format | Article |
| id | doaj-art-a98f55ce3fa34fbea87c2d48782bc8bd |
| institution | OA Journals |
| issn | 0736-6205 1940-9818 |
| language | English |
| publishDate | 1999-10-01 |
| publisher | Taylor & Francis Group |
| record_format | Article |
| series | BioTechniques |
| spelling | doaj-art-a98f55ce3fa34fbea87c2d48782bc8bd2025-08-20T02:25:35ZengTaylor & Francis GroupBioTechniques0736-62051940-98181999-10-0127474475210.2144/99274st05Quantification of 5-Methylcytosine in DNA by the Chloroacetaldehyde ReactionEdward J. Oakeley0Frédéric Schmitt1Jean-Pierre Jost21University of Exeter, Exeter, England, UK1University of Exeter, Exeter, England, UK1University of Exeter, Exeter, England, UKThe study of changes in genome-wide levels of DNA methylation has become a key focus for understanding the epigenetic regulation of gene expression. Many procedures exist to study DNA methylation, falling into two categories: gene-specific and genome-wide. Genome-wide methylation analysis is best performed by DNA hydrolysis followed by HPLC; however, it requires access to an HPLC machine, which is not always available. Alternative procedures, such as the radioactive labeling of CpG sites using SssI DNA methyltransferase, have been developed to address this problem, but it can only monitor CpG methylation changes, and CpNpG methylation is not detected. Here, we present a method for the analysis of DNA methylation in any sequence context by fluorescent labeling. We present control analyses using synthetic oligonucleotides of known methylation levels and a comparison of genomic DNA from two transgenic tobacco lines known to differ in their methylation levels. The results indicate that hygromycin-induced hypermethylation acts equally on all classes of methylatable cytosine, perhaps indicating a common mechanism.https://www.future-science.com/doi/10.2144/99274st05 |
| spellingShingle | Edward J. Oakeley Frédéric Schmitt Jean-Pierre Jost Quantification of 5-Methylcytosine in DNA by the Chloroacetaldehyde Reaction BioTechniques |
| title | Quantification of 5-Methylcytosine in DNA by the Chloroacetaldehyde Reaction |
| title_full | Quantification of 5-Methylcytosine in DNA by the Chloroacetaldehyde Reaction |
| title_fullStr | Quantification of 5-Methylcytosine in DNA by the Chloroacetaldehyde Reaction |
| title_full_unstemmed | Quantification of 5-Methylcytosine in DNA by the Chloroacetaldehyde Reaction |
| title_short | Quantification of 5-Methylcytosine in DNA by the Chloroacetaldehyde Reaction |
| title_sort | quantification of 5 methylcytosine in dna by the chloroacetaldehyde reaction |
| url | https://www.future-science.com/doi/10.2144/99274st05 |
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